Feasibility of expanding ischemia time for hearts destined for transplantation

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In collaboration with Prof. Brandacher at Johns Hopkins University we plan to improve the currently most used clinical method of heart preservation, namely hypothermia at 0 to +4°C. We will build upon our base preservation formulation, Unisol™, that has been shown to preserved whole large animals below +10°C for 6- 8h after total blood replacement with Unisol™ with normal functions upon return to physiological conditions. We have also shown that Unisol™ can maintain blood vessel function for at least 6 days at both -7° C and +4°C and that mouse hearts stored at +4°C for 18 hours in Unisol™ have a significantly faster return of heart function than hearts stored in the gold standard hypothermic heart preservation solution Celsior (HTK). Encouraged by these results we propose evaluation of Unisol supplemented with reagents targeting oxidation, apoptosis (enhancers of stress tolerance) and metabolism (metabolic rate inhibition) in 2 specific aims using a human cardiac myocyte cell line to select optimal reagent concentrations in vitro and a heterotopic heart transplant model to evaluate the best supplement formulations developed in the in vitro studies. The lead in vitro assay will be alamarBlue, however outcomes will be checked using alternative assays including trypan blue, live/dead stain and MTT assay. Apoptosis will be evaluated if significant losses of metabolic activity are observed 1 to 2 days after return to physiologic culture conditions. MALDI tissue imaging and proteomics will be performed on selected control and best performing hearts to provide molecular mapping information on metabolites, lipids, enzymatic products, and post translational modifications in heart sections. The in vivo studies will evaluate the best formulations from the in vitro studies over 7 days of hypothermic storage using a syngeneic BALB/C mouse, heterotopic transplantation model. Controls hearts will be preserved in high K+ Unisol™ (UHK) and HTK. We anticipate that the supplemented formulation will maintain viability and function of hearts for at least 36 hours, a considerable improvement over all current practice methods. Based upon preliminary data with blood vessel preservation, heart storage times of 6-7 days may be obtained. This innovation will not only increase storage time, it will also provide the opportunity for more closely matched recipients and potentially induction of tolerance. Furthermore, we are improving on the most tried and true method for hypothermic heart storage in clinical practice that is relatively inexpensive and easy to ship by air. Demonstration of ≥36 hours of hypothermic storage with retention of heart function will be considered to be a successful demonstration of feasibility for progression to a Phase II SBIR proposal for further evaluation in large animal models and ex vivo human heart evaluation post-preservation.